Development of an Enzyme-Linked Immunosorbent Assay for the Quantification of Amiodarone

A sensitive and specific enzyme-linked immunosorbent assay for an antiarrhythmic drug, amiodarone (AMI), was developed, which is capable of measuring levels as low as 16 ng/ml. Anti-AMI antibody was obtained by immunizing rabbits with an antigen conjugated with bovine serum albumin using diazotized 4-amino-1-(2-diethylaminoethoxy)-2,6-diiodobenzene. Enzyme labeling of AMI with β-D-galactosidase was similarly performed using a diazotized 4-amino-1-(2-diethylaminoethoxy)-2,6-diiodobenzene. This enzyme-linked immunosorbent assay was specific for AMI and showed a very slight cross-reactivity (1.25%) with its major metabolite, mono-N-desethylamiodarone. The values of the AMI concentrations measured by this assay were in good correlation to those by HPLC. Its analytical applicability was demonstrated by a kinetic study with human liver microsomes. The enzyme-linked immunosorbent assay should be a valuable tool in therapeutic drug monitoring and pharmacokinetic studies of AMI.